Herpes viruses are composed of a DNA core surrounded by a icosahedral capsid and a lipoprotein envelope. These viruses have a special affinity for cells of ectodermal origin and tend to produce latent infections.
The herpes simplex viruses (HSV), which cause infections in humans, are herpes simplex virus type (HSV1) which causes primarily oral lesions, keratitis and encephalitis; and herpes simplex virus type 2 (HSV2) which causes primarily genital lesions and life threatening disease to newborns.
Genital HSV infections are primarily caused by HSV2, however, HSV1 can also cause venereal disease. It is thought that herpes is one of the most common sexually transmitted diseases.
The initial HSV infection is characterized by moderate to severely painful lesions, which often take the form of tiny blister-like sores, that can progress to ulcers. Initial infection may also be accompanied by nausea, headaches, and fever. The HSV infection may remain latent in the host and cause frequent recurrent infections. Fifty percent of infected individuals have recurrent episodes, whereas the other 50% do not. At the present time, there is no effective cure for any of the HSV infections.
It is estimated that the number of individuals with venereal HSV infections in the United States ranges from 5-40 million, with 20 million being the most commonly quoted figure. It is further estimated that approximately 300-500 thousand individuals per year are infected by HSV2.
HSV infections are usually not incapacitating to the adult. However, HSV infection of the fetus or newborn infant is a severe disease with a high mortality rate and serious secondary sequelae.
Venereal HSV infections are usually diagnosed by the appearance of lesions, cellular morphology, as demonstrated by PAP or Tzanck smear, direct or indirect immunofluorescent assays, or viral infectivity in tissue culture. The majority of herpes lesions are being diagnosed by the doctor or clinician based on the appearance of the lesion only.
It is difficult to determine if a genital lesion is caused by HSV. It is particularly difficult to diagnose herpes infections in females, since the characteristic blisters often break, resulting in an ulcerative lesion that looks similar to many other types of venereal lesions.
The Tzanck smear is often used to diagnose venereal herpes. However, a large amount of false negatives may be obtained. Both histological preparations (the Tzanck or PAP smear) have to be interpreted by a trained pathologist. However, even then, there is still a 20% chance of a false negative result.
Direct or indirect immunofluorescence detects the presence of HSV antigen in infected cells. The ability to detect the HSV antigen depends primarily on the quality of the specimen. The test can be difficult to read and interpret, therefore, whenever a negative result is obtained, a tissue cell culture should be inoculated with the specimen for verification.
At the present time, the only sure way to diagnose HSV infections is by growing the sample in a tissue cell culture. Definitive identification is then accomplished by an immunological procedure such as neutralization of the antigen by specific antibody or by a demonstraton of HSV virus antigens by the use of a specific antibody labeled with fluorescein isothiocyanate. This is probably the most common method for HSV identification, when the cost of and time for the test is not a consideration.
It is therefore highly desirable to have a diagnostic test of choice for identification of venereal HSV infections which is highly specific, very sensitive, able to be performed quickly (much faster than the current 2-10 day period), and available at a reasonable cost.
Such a test will be extremely useful in the screening and monitoring of pregnant women, the diagnosis of the HSV infected population who have recurrent infections, the diagnosis of new cases and the screening of individuals with herpes-like infections.
Immunoassays employing various reading modes have been described in the art. For a number of reasons, including the ease of reading and the level of skill required by the operator, enzyme immunoassay is often a method of choice, although fluorescent or radioactive markers or labels are also frequently employed.
As to one embodiment of this invention, in the diagnosis of an HSV infection, a particular serologic marker is of importance. The presence of specific antibody of the IgM immunoglobulin indicates that the patient has recently experienced an acute or primary infection, whereas IgG class antibody is characteristically observed in the blood during convalescence, or after the recurrent infections. Thus, a method for detecting IgM antibody is desirable.
Dreesman et al, Intervirology, 12: 115-119 (1979) describe a solid phase radioimmunoassay for HSV which measures type specific antibody for both HSV1 and HSV2.
Hollinger et al, J. Immunol. 107: 1099, (1971) describe a solid phase radioimmunoassay for the detection of hepatitis B surface antigen.
Hevey et al., U.S. Pat. No. 4,228,237 describee an insoluble phase enzyme immunoassay which employs enzyme labeled avidin and biotin labeled reagent (e.g. antibody) to detect a ligand (e.g. an antigen).
Parikh, U.S. Pat. No. 4,298,685 describes a diagnostic system where the biotin-avidin system is used to detect antigen by fixing avidin to form an insoluble phase which precipitates the biotin labeled competitive reaction product.
Guesdon et al, J. Histochem. & Cytochem. 27: 1131, (1979) describe the use of biotin labeled antibody followed by the used of avidin labeled enzyme in an immunoassay. The reference likewise describes the use of avidin as a linking agent between biotin labeled antibody and biotin labeled enzyme.
Bayer et al, November 1978 TIBS pp. N257-N252 and Bayer et al, FEBS Letters October 1976 Vol. 68, No. 2, discuss the avidin biotin complex in Microbiology.
Wands et al, Proceedings in the National Academy of Sciences, U.S.A., Vol. 78, pp. 1214-1218 (1981), and U.S. Pat. No. 4,271,145 describe the use of high affinity IgM monoclonal antibody in the immunodiagnosis of hepatitis B virus.
Wisdom, Clin. Chem. Vol. 22 pp. 1243-1255 (1976) and Yolken, Rev. Infect. Diseases, 4: 35-68, (1982) discuss enzyme immunoassays in review articles.
A number of patents relating to enzyme immunoassays by Schuurs and co-workers is disclosed in U.S. Pat. Nos. 3,654,090, 3,791,932, 3,850,752, 3,839,153 and 3,879,262, 4,106,043 and Re 29,169.
The use of polyclonal antisera to characterize HSV1 and HSV2 has demonstrated extensive serological cross-reactivity between the two serotypes of virus. Hyperimmune sera produced against one HSV type will generally neutralize the heterologous type, although to a lesser degree. Using sensitive radioimmunoassays and experimentally produced monospecific antisera, both type-common and type-specific HSV antigenic determinants have been demonstrated. With the advent of procedures for producing monoclonal antibodies, specific determinants present as a single antigenic site could be characterized, thus facilitating the serological detection of HSV cross-reacting and type-specific antigens. Certain monoclonal antibodies against HSV-coded proteins have been described and have detected both type-common and type-specific HSV-antigenic determinants. However, prior to the invention herein, it is believed that monoclonal IgM antibody against HSV was unknown.